Significant progress was made during this reporting period. In collaboration with Dr. Blough, we continued to evaluate a large number of newly synthesized compounds using in vitro assays. The current focus is on DA/5-HT releasers based on the phenmetrazine structure, since these compounds have minimal activity at the 5-HT2B receptor. A key aspect of this project is to avoid making compounds that activate 5-HT2B receptors, since this property is associated with the development of cardiac valve disease in humans. We published In collaboration with Dr. Ananthan, who received extramural NIDA funding for further identification of dopamine transporter allosteric modulator ligands, we commenced our part of this collaboration. Five papers related to this project were published in the reporting period, two of which will be highlighted here because of their direct relevance to medication development efforts. Paper #1: In vivo effects of amphetamine analogs reveal evidence for serotonergic inhibition of mesolimbic dopamine transmission in the rat. J Pharmacol Exp Ther. 2011 Apr;337(1):218-25. Evidence suggests that elevations in extracellular serotonin (5-HT) in the brain can diminish stimulant effects of dopamine (DA). To assess this proposal, we evaluated the pharmacology of amphetamine analogs (m-fluoroamphetamine, p-fluoroamphetamine, m-methylamphetamine, p-methylamphetamine), which display similar in vitro potency as DA releasers (EC(50) = 24-52 nM) but differ in potency as 5-HT releasers (EC(50) = 53-1937 nM). In vivo microdialysis was used to assess the effects of drugs on extracellular DA and 5-HT in rat nucleus accumbens, while simultaneously measuring ambulation (i.e., forward locomotion) and stereotypy (i.e., repetitive movements). Rats received two intravenous injections of drug, 1 mg/kg at time 0 followed by 3 mg/kg 60 min later. All analogs produced dose-related increases in dialysate DA and 5-HT. All analogs increased ambulation and stereotypy, but drugs causing greater 5-HT release (e.g., p-methylamphetamine) were associated with significantly less forward locomotion. The magnitude of ambulation was positively correlated with extracellular DA (p <0.001) and less so with the ratio of DA release to 5-HT release (i.e., percentage DA increase divided by percentage 5-HT increase) (p <0.029). Collectively, our findings are consistent with the hypothesis that 5-HT release dampens stimulant effects of amphetamine-type drugs, but further studies are required to address the precise mechanisms underlying this phenomenon. Paper #2: Serotonin (5-HT) precursor loading with 5-hydroxy-l-tryptophan (5-HTP) reduces locomotor activation produced by (+)-amphetamine in the rat. Drug Alcohol Depend. 2011 Apr 1;114(2-3):147-52. Background, Evidence suggests that increases in synaptic serotonin (5-HT) can reduce the stimulant properties of amphetamine-type drugs. Here we tested the hypothesis that administration of the 5-HT precursor 5-hydroxy-l-tryptophan (5-HTP), along with the peripheral decarboxylase inhibitor benserazide, would decrease locomotor effects of (+)-amphetamine. METHODS: Drug treatments were administered to conscious male rats undergoing in vivo microdialysis in nucleus accumbens. During dialysis sampling, rats were housed in chambers equipped with photobeams to detect forward locomotion (i.e., ambulation) and repetitive movements (i.e., stereotypy). Extracellular concentrations of dopamine (DA) and 5-HT were measured by high-pressure liquid chromatography with electrochemical detection.RESULTS:5-HTP (10 &30 mg/kg, i.p.) plus benserazide (30 mg/kg, i.p.) caused dose-related increases in 5-HT but failed to alter other parameters. (+)-Amphetamine (0.3 &1.0 mg/kg, i.p.) produced dose-related increases in DA, ambulation and stereotypy. Combined administration of 5-HTP and (+)-amphetamine evoked large elevations in extracellular DA and 5-HT, but caused significantly less ambulation than (+)-amphetamine alone (50% reduction).CONCLUSIONS:Our results confirm that 5-HTP can decrease hyperactivity produced by (+)-amphetamine, even in the presence of elevations in dialysate DA. The data suggest that 5-HTP and (+)-amphetamine may be useful to broadly enhance monoamine function in the clinical setting, while reducing undesirable effects of (+)-amphetamine.